Quantification of Wolbachia copy number in Trichogramma eggs (Hymenoptera: Trichogrammatidae): Lysozyme treatment significantly improves total gene yield from the Gram-negative bacterium

The Wolbachia bacterium is one of the most prevalent endosymbionts of arthropods. The bacterium is known to induce four distinct reproductive manipulations in its various hosts; namely, cytoplasmic incompatibility, feminization, male killing and parthenogenesis induction. The bacterial titer has been thought to have an important role in expression of these phenotypes. Although host eggs of the bacterium are the place where all of the manipulations take place, in most cases, they are not big enough to be subjected to mechanical disruption to extract genetic materials from them. Furthermore, DNA extraction protocols generally involve washing steps in which part of the DNA is lost. With the simple three step digestion protocol we developed, we were able to preserve the whole genome and measure the titer of Wolbachia in individual Trichogramma eggs by real-time quantitative polymerase chain reaction (PCR), which was comparable to the titer by visual observation.